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Lionel Bretillon, Emilie Simon, Christine Arnould, Géraldine Lucchi, Delphine Pecqueur, Caroline Truntzer, Niyazi Acar, Jeannine Lherminier, Patrick Ducoroy, Catherine P Garcher; Proteomic analysis of sub-retinal deposits and RPE of the ApoB100,LDLR-/- mouse, a murine model of aging of the human retina. Invest. Ophthalmol. Vis. Sci. 2014;55(13):1316.
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The deposition of basal deposits and drusen at the basement of the retinal pigment epithelium (RPE) and within Bruch’s Membrane (BrM) is a hallmark of aging and is enhanced in Age-related Macular Degeneration (AMD). The question of the origin of the deposits and drusen remains largely unresolved. Part of the lipid content of drusen and basal deposits originates from the choroid. Recently, sub-retinal drusenoid deposits (SDD) were characterized in AMD eyes. Data from a topographical study in human eyes suggest that rods may be the main source of SDD whereas basal deposits may originate from cones. To improve our knowledge on the pathophysiology of aging and mechanisms of lipid deposition, we characterized the proteomic changes in the retina and RPE of the ApoB100,LDLR-/- mouse, a relevant model of aging of the human retina.
Ocular globes of aged ApoB100,LDLR-/- and control mice were embedded in OCT and cryosectioned in 10µm-thick slices. The outer segments of the photoreceptors, SDD and RPE samples were collected by Laser Capture Microdissection. Proteins were extracted and purified by 1D gel electrophoresis. After trypsin digestion, peptides were analyzed by nanoLC-MS/MS using an LTQ-Orbitrap Elite mass spectrometer. The proteins were identified using a combination of X!Tandem and Mascot, and then validated using PeptideProphet and ProteinProphet. Data were processed and quantified using a set of home-made and open-source software tools. Differential proteins were selected using the MSstats library on R-software with a 5% FDR, and searched against Kegg and Reactome pathways and Gene Ontology terms databases using the Cytoscape software.
From 38 to 96 proteins were identified in the different samples. Lower levels of retinol dehydrogenase and up-regulation of S-arrestin in ApoB100,LDLR-/- mice compared to controls may explain reduced electroretinographic response in the transgenic mouse (impaired retinoid cycle). Up-regulation of crystallins in the transgenic mouse (in outer segments, sub-retinal deposits and RPE) is consistent with proteomic data in eyes collected from patients with AMD showing increased levels of crystallins αA and αB.
The proteomic data are consistent with findings in AMD patients and may add possible clues to explain functional changes in the ApoB100,LDLR-/- mouse (effect on S-arrestin and retinol dehydrogenase).
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